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|Title: ||Investigation of the transcriptional response of Sulfolobus solfataricus to damaging agents|
|Authors: ||Munro, Stacey|
|Supervisors: ||White, Malcolm F.|
|Issue Date: ||2009|
|Abstract: ||It is vital for the survival of an organism that it can repair damage to its DNA.
Exogenous and endogenous sources of damage are dealt with by a variety of repair
pathways that have evolved to repair specific types of damage. Organisms in the
archaeal domain, the third domain of life, contain homologues of many of the
eukaryotic repair proteins, however little is known about how damage is detected in
the archaeal domain.
Microarray studies in the archaeal species Sulfolobus solfataricus determined
a number of genes whose expression was effected by UV radiation (work by Dr D
Götz). The change in expression of nine of these genes was confirmed by RT real
time PCR. The expression of these genes was then investigated after exposure to
different damaging agents, Mitomycin C, Methyl methane sulfonate, Phleomycin and
Hydrogen peroxide. The expression of two genes, transcription factor tfb-3 and cell
division control gene cdc6-2, was up regulated in all damage conditions.
There was a huge induction of the dps-like gene (sso2079) after hydrogen
peroxide damage. Transcription from this genes promoter was shown to be strong in
vitro (work by Dr S Paytubi) suggesting a repressor was controlling the gene in vivo.
A palindromic repeat in the promoter of the dps-like gene was used to ‘fish’ for a
transcriptional repressor and the Sso2273 protein, a homologue of the diphtheria toxin
repressor (DtxR) from Corynebacterium diphtheria, was identified as a possible
Sso2273 was expressed and purified, and its crystal structure solved, its
paralogue, Sso0669, was also expressed and purified. Electrophoretic mobility shift
assays showed that the Sso2273 protein does not bind DNA, and had no effect on
transcription from any promoter used in in vitro transcription assays. However
Sso0669 appeared to inhibit transcription, although the inhibition was not sequence
A knockout strain of S. solfataricus PBL2025 missing the sso2273 gene was
produced and used in microarray experiments in an attempt to determine the role of
Sso2273 within the cell. The absence of Sso2273 appeared to have no effect on the
expression of the dps-like gene, however strong repression of an operon containing
genes involved in Sulphur assimilation was observed.|
|Other Identifiers: ||uk.bl.ethos.552274|
|Publisher: ||University of St Andrews|
|Appears in Collections:||Biology Theses|
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